This research project is directed towards the genetic and biochemical characterization of lambda DNA replication. Attempts to identify phage and host genes and their products involved in initiation of replication are being made. Various lambda DNA substrates and cofactors are being used in an invitro system capable of synthesizing DNA on phi X174 single strand circular templates in an attempt to detect phage O and P gene specific stimulation of lambda DNA replication. In vivo experiments have identified two phage genes and three bacterial genes involved in replication of lambda DNA. Further experiments are in progress to determine at what stage (initiation, "early", or "late" replication) these gene products act. Lambda DNA circles in lysogens undergo two types of nicking following inactivation of repressor. Genetic and biochemical evidence indicate that one type of nick is associated with bacterial membrane. Alteration in the membrane causes this type nick to disappear presumably through some sort of repair process. Alterations produced in the membrane of E. coli through mutational chemical and biological means are being used to investigate this phenomena.